Women have more difficulty quitting smoking and have higher rates of relapse. Evidence indicates that female smoking behavior is more likely to be motivated by affect regulation and stress, whereas male smoking behavior is more likely to be motivated by nicotine-related positive reinforcement. Pharmacotherapy development targeted toward sex-dependent aspects of smoking is a critical, yet underdeveloped area of research. Guanfacine is an adrenergic receptor agonist which reduces noradrenergic activity through stimulation of inhibitory presynaptic ?2a receptors. Guanfacine attenuates stress-induced reinstatement to drugs of abuse in preclinical studies and attenuate dopamine release in accumbens and pre-frontal cortex. In human laboratory studies, we have shown that guanfacine operates through sex-dependent mechanisms. Guanfacine attenuated stress reactivity in women and smoking-related positive reinforcement in men. In a randomized clinical trial, we found that guanfacine significantly increased smoking cessation (42% vs. 19% placebo; end-of-treatment point prevalence), and that effects were more pronounced in women (45% vs. 15%) versus men (39% vs. 23%). We have also completed three meta-analytic studies summarizing data from 29,005 smokers, finding that varenicline is more efficacious for women, compared to men, which is notable as nicotine replacement and bupropion are less efficacious for women. With regards to mechanisms, varenicline attenuates nicotine-based reinforcement, and withdrawal related effects on negative mood and cognition but there has been little investigation regarding sex differences in these effects. To date, guanfacine and varenicline are the only two smoking cessation medications which either produce equal or preferential rates of quitting in women. While guanfacine targets the noradrenergic system and varenicline targets the cholinergic system, both medications appear to target aspects of negative affect and nicotine-based reinforcement. There is significant preclinical data identifying that the noradrenergic and acetylcholine systems interact at the cellular and molecular level, and combined treatment with guanfacine and varenicline may produce additive effects on smoking cessation, particularly for women. For this proof of concept study, we plan to conduct a mechanistic Phase 2 double-blind, factorial design study to examine sex differences in the combined effect of guanfacine + varenicline on 1) counteracting the effects of stress-induced smoking behavior and smoking-related positive reinforcement in the laboratory and 2) improving abstinence outcomes during a subsequent treatment phase. Importantly, we will examine the safety of combining guanfacine with varenicline, and potential sex differences in mechanisms underlying smoking (e.g., craving, subjective cigarette effects, negative mood, withdrawal, cognitive function, cardiovascular reactivity, and HPA-axis levels). To our knowledge, this will be the first study investigating sex differences in the therapeutic potential and associated mechanisms of a selective ?2a agonist, guanfacine, combined with a nAChR partial agonist, varenicline for smoking cessation.